Genotyping and other DNA analyses using PCR or next-generation sequencing (NGS) require a suitable amount of high-quality
genomic DNA as starting material. The most common bottleneck to obtaining high-quality DNA is tissue lysis. Fibrous or
other hard-to-lyse tissues require lengthy and tedious protocols that often result in low yields of low-quality DNA. The
QIAamp Fast DNA Tissue Kit, based on proven QIAGEN technologies, overcomes this bottleneck by providing innovative
disruption tubes that significantly improve yield and shorten the overall time to result. The procedure can be fully automated on theQIAcube.

The QIAamp Fast DNA Tissue Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

Inhibitors are efficiently removed

Internal controls
were amplified in the presence of DNA isolated from 10 mg
porcine ear-punches using the QIAamp Fast DNA Tissue Kit or
QIAamp DNA Mini Kit. When 1 or 6 μl of eluate were spiked
in, amplification of the internal control was unaffected, showing
that no inhibitors were present in the eluates.
IC: Internal Control.

PFGE analysis

Rat kidney tissue was used
to isolate genomic DNA
using different protocols.
A Clamped Homogeneous
Electric Field (CHEF) was
used to analyze the integrity
of gDNA obtained using
different protocols. A 1%
agarose gel was run with
0.5x TBE with 6V/cm and
switch time of 1–12 s. The
run time was 16 h at 11°C.
Equal amounts of DNA
were loaded on the gel.

Time is saved with the QIAamp Fast DNA Tissue Kit compared to standard methods

Yield analysis of genomic DNA extracted using different protocols

gDNA was isolated from 10 mg of rat liver
tissue using different protocols. For each sample, 8 μl was loaded on a 0.7% TAE gel. The gel was run for 16 h at 25 V.
Ladder = Lambda HindIII.

Performance

The QIAamp Fast DNA Tissue Kit uses a novel combination of mechanical, chemical and enzymatic lysis. Each kit is supplied with disruption tubes that contain a special stainless steel bead with a unique shape that aids tissue disruption more thoroughly than other methods. Firstly, the tissue sample and reagent mix are added to the tubes and homogenized using a common benchtop vortexer (with appropriate adapters) or in a bead mill, such as the TissueLyser II or LT, followed by a short lysis in the same tube. DNA is then extracted from the lysate using proven QIAamp technology. The pretreatment reduces the total DNA extraction time from several hours to just 30 minutes.

As well as speeding up the total time to extract DNA, the mechanical disruption step and thorough lysis protocol increase DNA yield. Improvements in yield from different tissue types have been evaluated using the QIAxpert and a fluorometric assay. The results show that the QIAamp Fast DNA Tissue Kit is capable of extracting up to tenfold more DNA compared to standard methods, while also providing one of the fastest protocols.

Most NGS applications require high-molecular-weight DNA (>10 kb) to allow for larger spans to be sequenced and assembled with bioinformatic tools. Some disruption methods may fragment the DNA through shearing, which can affect downstream analysis. Compared to other kits that show high fragmentation down to a few kb, the QIAamp Fast DNA Tissue Kit allows you to obtain DNA between 25 and 50 kb and is therefore very well suited to any PCR or NGS application.

Principle

Molecular techniques such as PCR and NGS are revealing previously unknown details from complex experimental samples. To achieve reliable results using these techniques it is essential to obtain sufficient amounts of high-quality DNA. This can be challenging when working with fibrous or hard tissues because they are difficult to lyse, which directly affects the amount of DNA that is released. To address this, our new QIAamp Fast DNA Tissue Kit combines mechanical, chemical and enzymatic tissue lysis, which improves DNA yields by 5- to 10-fold compared to standard methods. The pretreatment reduces the total DNA extraction time from several hours to just 30 minutes, so in addition to achieving high-integrity DNA suitable for PCR and NGS applications, you also get significant time savings.

Procedure

The QIAamp Fast DNA Tissue Kit uses a combination of mechanical, chemical and enzymatic lysis to homogenize samples. The included Tissue Disruption Tubes contain a specially shaped bead that effectively disrupts tissue when agitated. As little as 5 minutes on a desktop vortexer or 30 seconds in a high-powered bead mill is sufficient to homogenize the tissue. The optimized chemistry allows homogenization of the tissue and simultaneous stabilization of DNA released from the disrupted tissue. The digestion buffer mix also contains proteinase K, which completely lyses the sample material in a short subsequent incubation and is not affected by mechanical homogenization. Genomic DNA is subsequently purified using proven QIAamp technology, delivering pure DNA for any downstream application like PCR or sequencing. Additionally, the QIAamp Fast DNA Tissue Kit can be conveniently used with the QIAcube for automated sample preparation.

Applications

The QIAamp Fast DNA Tissue Kit delivers high yields of pure DNA from any tissue type and is suitable for PCR and NGS applications.